Role of activating protein-1 in the regulation of the vascular cell adhesion molecule-1 gene expression by tumor necrosis factor-alpha.
Endothelial cell surface expression of VCAM-1 is one of the initial steps in the pathogenesis of atherosclerosis. The inflammatory response transcription factor nuclear factor (NF)-kappaB plays an important role in the regulation of VCAM-1 expression by various stimuli including tumor necrosis factor (TNF)-alpha. Other transcription factors may modulate this response through interaction with NF-kappaB factors. Since c-Fos/c-Jun (activating protein-1 (AP-1)) are expressed in vascular endothelium during proinflammatory conditions, we investigated the role of AP-1 proteins in the expression of VCAM-1 by TNF-alpha in SV40 immortalized human microvascular endothelial cells (HMEC). TNF-alpha induced expression of both early protooncogenes, c-fos and c-jun. The ability of TNF-alpha to activate the kappaB-motif (kappaL-kappaR)-dependent VCAM-1 promoter-chloramphenicol acetyltransferase (CAT) reporter gene lacking a consensus AP-1 element was markedly inhibited by co-transfection of the expression vector encoding c-fos ribozyme, which decreases the level of c-fos by degrading c-fos mRNA, or c-fos or c-jun oligonucleotides. Conversely, co-transfection of c-Fos and c-Jun encoding expression vectors potentiated the p65/NF-kappaB-mediated transactivation of the VCAM-1 promoter-CAT reporter gene. Furthermore the c-Fos encoding expression vector potentiated by 2-fold the transactivation activity of a chimeric transcriptional factor Gal/p65 (containing the transactivation domain of p65 and the DNA binding domain of the yeast transcriptional factor Gal-4). Consistent with the promoter studies, curcumin and NDGA, inhibitors of AP-1 activation, markedly inhibited the ability of TNF-alpha to activate the expression of VCAM-1 mRNA levels at concentrations that did not inhibit the activation of NF-kappaB. In gel mobility supershift assays, the antibodies to c-Fos or c-Jun inhibited the binding of TNF-alpha-activated nuclear NF-kappaB to the kappaL-kappaR, suggesting that both c-Fos and c-Jun interacted with NF-kappaB. These results suggest that AP-1 proteins may mediate the effect of TNF-alpha in the regulation of VCAM-1 expression through interaction with NF-kappaB factors in endothelial cells.
Ahmad
M
M
Division of Cardiology, Department of Medicine, Emory University School of Medicine, Atlanta, Georgia 30322, USA. mahmad@emory.edu
Theofanidis
P
P
Medford
R M
RM
D002461:Cell Line, Transformed; D003474:Curcumin / Q000494:pharmacology; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation / Q000187:drug effects; D005822:Genetic Vectors; D006801:Humans; D009637:Masoprocol / Q000494:pharmacology; D016328:NF-kappa B / Q000378:metabolism; D016755:Proto-Oncogene Proteins c-jun / Q000096:biosynthesis / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D019010:Vascular Cell Adhesion Molecule-1 / Q000235:genetics*
D016328:NF-kappa B; D016755:Proto-Oncogene Proteins c-jun; D014409:Tumor Necrosis Factor-alpha; D019010:Vascular Cell Adhesion Molecule-1; D009637:Masoprocol; D003474:Curcumin